title: Activation of Dbl restores migration in polyamine-depleted intestinal epithelial cells via Rho-GTPases.
abstract: Integrin binding to the extracellular matrix (ECM) activated Rho GTPases, Src, and focal adhesion kinase in intestinal epithelial cells (IEC)-6. Polyamine depletion inhibited activities of Rac1, RhoA, and Cdc42 and thereby migration. However, constitutively active (CA) Rac1 expression abolished the inhibitory effect of polyamine depletion, indicating that polyamines are involved in a process upstream of Rac1. In the present study, we examined the role of polyamines in the regulation of the guanine nucleotide exchange factor, diffuse B-cell lymphoma (Dbl), for Rho GTPases. Polyamine depletion decreased the level as well as the activation of Dbl protein. Dbl knockdown by siRNA altered cytoskeletal structure and decreased Rac1 activity and migration. Cells expressing CA-Dbl increased migration, Rac1 activity, and proliferation. CA-Dbl restored migration in polyamine-depleted cells by activating RhoA, Rac1, and Cdc42. CA-Dbl caused extensive reorganization of the F-actin cortex into stress fibers. Inhibition of Rac1 by NSC23766 significantly decreased migration of vector-transfected cells and CA-Dbl-transfected cells. However, the inhibition of migration was significantly higher in the vector-transfected cells compared with that seen in the CA-Dbl-transfected cells. Dbl localized in the perinuclear region in polyamine-depleted cells, whereas it localized with the stress fibers in control cells. CA-Dbl localized with stress fibers in both the control and polyamine-depleted cells. These results suggest that polyamines regulate the activation of Dbl, a membrane-proximal process upstream of Rac1.
authors.affiliation: Department of Physiology, University of Tennessee Health Science Center, Memphis, 38163, USA. rray3@uthsc.edu
authors.forename: Ramesh M
authors.initials: RM
authors.lastname: Ray
authors.forename: Mitulkumar N
authors.initials: MN
authors.lastname: Bavaria
authors.forename: Sujoy
authors.initials: S
authors.lastname: Bhattacharya
authors.forename: Leonard R
authors.initials: LR
authors.lastname: Johnson
mesh_terms: D000634:Aminoquinolines / Q000494:pharmacology; D000818:Animals; D002460:Cell Line; D002465:Cell Movement; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004847:Epithelial Cells / Q000201:enzymology*; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics / Q000378:metabolism*; D007413:Intestinal Mucosa / Q000201:enzymology*; D011073:Polyamines / Q000378:metabolism*; D011743:Pyrimidines / Q000494:pharmacology; D034622:RNA Interference; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D022502:Stress Fibers / Q000378:metabolism; D014162:Transfection; D020764:cdc42 GTP-Binding Protein / Q000378:metabolism; D054462:p21-Activated Kinases / Q000378:metabolism; D020830:rac1 GTP-Binding Protein / Q000037:antagonists & inhibitors / Q000378:metabolism; D020741:rho GTP-Binding Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D020742:rhoA GTP-Binding Protein / Q000378:metabolism
chemical_list: D000634:Aminoquinolines; C560313:Dbl protein, rat; D004791:Enzyme Inhibitors; D020662:Guanine Nucleotide Exchange Factors; C487513:NSC 23766; D011073:Polyamines; D011743:Pyrimidines; D011993:Recombinant Fusion Proteins; D054462:p21-Activated Kinases; C499134:Rac1 protein, rat; D020764:cdc42 GTP-Binding Protein; D020830:rac1 GTP-Binding Protein; D020741:rho GTP-Binding Proteins; D020742:rhoA GTP-Binding Protein